I have observed the induction by mutagen treatment of colonies of Chinese hamster ovary (CHO) cells which are sectored (mosaic) in appearance when stained for glucose-6-phosphate dehydrogenase (G6PD) activity. This phenomenon occurred when populations of single CHO cells were exposed to the mutagen ethyl methanesulfonate, (EMS) allowed to grow into colonies and stained histochemically for G6PD activity. Colonies composed of unstained and stained cells in various sectored patterns and uniformly unstained colonies (pure mutant) were observed at a frequency of about 1 per 1000 colonies. Although the production of sectored colonies after exposure of single cell to mutagenic agents has been observed in yeast and bacteria, this is a new phenomenon for a somatic mammalian cell line. I propose to investigate the basis of this phenomenon and use it as a tool to study the process of mutagenesis in mammalian cells. The influence of the replication state of the G6PD gene on the sectoring process will be examined using synchronized cell populations which are exposed to the mutagen in various parts of the cell cycle. The possibility that pure mutant clones are caused by a lethal hit on the daughter DNA strand will be investigated by examining the mosaic: pure mutant ratio at various levels of survival to the mutagen. Spontaneous reversion rates for G6PD negative mutants derived from mosaic and pure mutant clones will be analyzed to determine whether pure mutant clones are due to deletion events. The relationship of DNA repair to sectoring will be investigated by examining this phenomenon in mutagen sensitive CHO strains and using drugs such as caffeine which inhibits post-replication DNA repair. A variety of mutagens such as X-rays, UV-light, chemical agents (EMS, nitrosoguanidine, 2-amino-purine) will be examine for their ability to produce sectored and pure mutant colonies. The possibility that the phenomenon of mutagen induced replicating instabilities observed in yeast and Drosophila also occur in the CHO cell will be determined.